1. Field of the Invention
The invention relates to two Lactobacillus isolates having anti-inflammatory activities and beneficial probiotic properties, namely Lactobacillus sakei GMNL-76 and Lactobacillus reuteri GMNL-89, which were deposited in the Biosource Collection and Research Center (BCRC) of the Food Industry Research and Development Institute (FIRDI) under accession numbers BCRC 910355 and BCRC 910340, respectively, and in the China Center for Type Culture collection (CCTCC) under accession numbers CCTCC M 207153 and CCTCC M 207154, respectively. The two isolates and their sub-cultured offspring can be used in the preparation of a variety of food products, and in the manufacture of pharmaceutical compositions for treating and/or alleviating diseases associated with inflammation, such as rheumatoid arthritis.
2. Description of the Related Art
Cytokines are known to be involved in numerous important biological processes, including inflammation, tissue repair, cell growth, fibrosis, angiogenesis, and immune response. Therefore, cytokines play an important role in autoimmune diseases.
M. Feldmann et al. investigated the pathogenesis of rheumatoid arthritis by analyzing cytokine expression and regulation in rheumatoid arthritis synovial tissue (M. Feldmann et al. (1996), Annu. Rev. Immunol., 14:397-440). According to M. Feldmann et al., cytokines are divided into four main classes: (1) proinflammatory cytokines; (2) immunoregulatory cytokines; (3) chemotactic cytokines; and (4) mitogenic cytokines.
Proinflammatory cytokines are a group of molecules produced by T-helper 1 cells (Th1 cells for short) and having the ability to regulate delayed-type hypersensitivity reaction, including interleukin-1 (IL-1), interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and granulocyte-macrophage colony stimulating factor (GM-CSF). Cartilage destruction observed in rheumatic diseases has been widely recognized to have been caused by activity of matrix metalloproteinases (MMPs). MMPs are generated by activated macrophages and fibroblasts that are responsive to proinflammatory cytokines (such as IL-1 or TNF-α). M. Feldmann et al. further reported that injection of IL-1 or TNF-α into collagen-immunized mice or rats, or local injection of IFN-γ into footpads of collagen type II-immunized mice would promote the incidence of rheumatoid arthritis and exacerbates the disease (M. Feldmann et al. (1996), supra).
Immunoregulatory cytokines (also referred to as anti-inflammatory cytokines) are a group of molecules generated by T-helper 2 cells (Th2 cells for short) and having the ability to inhibit inflammatory reaction, including IL-4, IL-10, IL-13, and Transforming Growth Factor-β (TGF-β). In an article by G. Garcia et al. (G. Garcia et al. (1999), Journal of Autoimmunity, 13:315-324), it is reported that the two immunoregulatory cytokines, TGF-β and IL-10, not only inhibit production of proinflammatory cytokines that will induce MMPs, they will also induce production of natural inhibitors of MMPs (i.e., tissue inhibitors of matrix metalloproteinases, TIMPs). G. Garcia et al. further pointed out that IL-10 was known to have the ability to inhibit production of IFN-γ from Th1 cells and production of a variety of cytokines from other leukocyte populations. IL-10 inhibited production of IL-1, IL-6, TNF-α, and IL-8 and G-CSF (granulocyte-colony stimulating factor) from macrophages, and production of IL-1, TNF, IL-8, macrophage inflammatory protein 1α (MIP1α), and macrophage inflammatory protein 1β (MIP1β) from polymorphonuclear cells. Most of these cytokines and chemokines are associated with the pathological process of rheumatoid arthritis (G. Garcia et al. (1999), supra).
These study results reveal that proinflammatory cytokines (such as TNF-α and IFN-γ) associated with Th1 cells promote inflammation and aggravate rheumatoid arthritis, whereas immunoregulatory cytokines (such as IL-10 and IL-4) associated with Th2 cells can not only inhibit production of proinflammatory cytokines but also induce production of TIMPs to thereby reduce cartilage destruction.
Jae-Seon So reported that oral administration of Lactobacillus casei suppressed collagen-induced arthritis (CIA) and reduced paw swelling, lymphocyte infiltration, and destruction of cartilage tissue. Lactobacillus casei administration reduced type II collagen (CII)-reactive proinflammatory molecules (IL-1β, IL-2, IL-6, IL-12, IL-17, IFN-γ, TNF-α, and Cox-2) by CD4+ T cells. Lactobacillus casei administration also reduced translocation of NF-κB into nucleus and CII-reactive Th1-type IgG isotypes IgG2a and IgG2b, while up-regulating IL-10 levels (Jae-Seon So et al. (2008), Mol. Immunol., 45(9):2690-2699; Epub Feb. 19, 2008).
If rheumatoid arthritis can be treated/alleviated through administration of Lactobacillus isolates, a safe and inexpensive drug might be developed for patients with rheumatoid arthritis.
To achieve the aforesaid objective, the Applicants have screened two Lactobacillus isolates from gastrointestinal tract specimens of adult subjects from Taiwan. The two Lactobacillus isolates are phylogenetically different from known strains of their respective species, and possess anti-inflammatory activities that can stimulate generation of large amounts of IL-10 and relatively small amounts of IFN-γ and/or TNF-α. Therefore, these Lactobacillus isolates are anticipated to be useful in treating diseases associated with inflammation, including, but not limited to, rheumatoid arthritis.